Tachyphylaxis and reproducibility of desmopressin response in perioperative persons with nonsevere hemophilia A: implications for clinical practice.

Background: Desmopressin is frequently used perioperatively in persons with nonsevere hemophilia A. However, increase in factor (F)VIII:C after desmopressin use is interindividually highly variable. Tachyphylaxis has only been reported in test setting for persons with hemophilia A, with a remaining response of approximately 70% after a second dose compared with that after a first dose. Objectives: To study tachyphylaxis of FVIII:C response after multiple administration(s) of desmopressin in perioperative persons with nonsevere hemophilia A. Methods: We studied FVIII:C levels after desmopressin before (day 0 [D0]) and on days 1 (D1) and 2 (D2) after surgery in 26 patients of the DAVID and Little DAVID studies. We studied tachyphylaxis by comparing the responses at D1 and D2 with that at D0. We also assessed the reproducibility of the D0 response in comparison to an earlier performed desmopressin test. Results: The median absolute FVIII:C increase was 0.50 IU/mL (0.35-0.74; n = 23) at D0, 0.21 IU/mL (0.14-0.28; n = 17) at D1, and 0.23 IU/mL (0.16-0.30; n = 11) at D2. The median percentage of FVIII increase after the second administration (D1) compared with the first (D0) was 42.9% (29.2%-52.5%; n = 17) and that of the third (D2) compared with the first (D0) was 36.4% (23.7%-46.9%; n = 11). The FVIII:C desmopressin response at D0 was comparable with the desmopressin test response in 74% of the patients. Conclusion: Tachyphylaxis in the surgical setting was considerably more pronounced than previously reported, with FVIII:C at D1 and D2 of 36% to 43% of the initial response. Our results may have important implications for monitoring repeated desmopressin treatment when used perioperatively.

Maternal and neonatal bleeding complications in relation to peripartum management in hemophilia carriers: A systematic review.

Currently, there is no consensus on the optimal management to prevent postpartum hemorrhage (PPH) in hemophilia carriers. We aimed to evaluate peripartum management strategies in relation to maternal and neonatal bleeding outcomes by performing an extensive database search up to August 2020. Seventeen case-reports/series and 11 cohort studies were identified of overall 'poor' quality describing 502 deliveries. The PPH incidence in the individual patient data was 63%; 44% for those women receiving prophylaxis to correct coagulation and 77% for those without (OR 0.23, CI 0.09-0.58) and in cohort data 20.3% (26.8% (11/41) vs. 19.4% (55/284) (OR: 1.53, 95% CI: 0.72-3.24), respectively. Peripartum management strategies mostly consisted of clotting factor concentrates, rarely of desmopressin or plasma. Tranexamic acid appears promising in preventing secondary PPH, but was not used consistently. Neonatal bleeding was described in 6 affected male neonates, mostly after instrumental delivery or emergency CS, but insufficient information was provided to reliably investigate neonatal outcome in relation to management. The high PPH risk seems apparent, at most mildly attenuated by prophylactic treatment. Prospective cohort studies are needed to determine the optimal perinatal management in hemophilia.

Clinical and computed tomography characteristics of COVID-19 associated acute pulmonary embolism: A different phenotype of thrombotic disease?

INTRODUCTION: COVID-19 infections are associated with a high prevalence of venous thromboembolism, particularly pulmonary embolism (PE). It is suggested that COVID-19 associated PE represents in situ immunothrombosis rather than venous thromboembolism, although the origin of thrombotic lesions in COVID-19 patients remains largely unknown. METHODS: In this study, we assessed the clinical and computed tomography (CT) characteristics of PE in 23 consecutive patients with COVID-19 pneumonia and compared these to those of 100 consecutive control patients diagnosed with acute PE before the COVID-19 outbreak. Specifically, RV/LV diameter ratio, pulmonary artery trunk diameter and total thrombus load (according to Qanadli score) were measured and compared. RESULTS: We observed that all thrombotic lesions in COVID-19 patients were found to be in lung parenchyma affected by COVID-19. Also, the thrombus load was lower in COVID-19 patients (Qanadli score -8%, 95% confidence interval [95%CI] -16 to -0.36%) as was the prevalence of the most proximal PE in the main/lobar pulmonary artery (17% versus 47%; -30%, 95%CI -44% to -8.2). Moreover, the mean RV/LV ratio (mean difference -0.23, 95%CI -0.39 to -0.07) and the prevalence of RV/LV ratio >1.0 (prevalence difference -23%, 95%CI -41 to -0.86%) were lower in the COVID-19 patients. CONCLUSION: Our findings therefore suggest that the phenotype of COVID-19 associated PE indeed differs from PE in patients without COVID-19, fuelling the discussion on its pathophysiology.

Maternal and neonatal bleeding complications in relation to peripartum management in women with Von Willebrand disease: A systematic review.

Women with Von Willebrand disease (VWD) have an increased risk of developing postpartum hemorrhage (PPH). Our aim is to evaluate peripartum management strategies in relation to maternal and neonatal bleeding complications in VWD. Electronic databases were searched up to January 2019. Seventy-one case-reports and -series and 16 cohort studies were selected, including 811 deliveries. Cohort studies reported primary PPH in 32% and secondary PPH in 13% of the women. The overall primary PPH incidence in the individual patient data was 34%, similar between women who received prophylactic treatment to prevent PPH and those who didn't. Neonatal bleeding events were reported in 4.6% of deliveries. Overall, the available evidence on peripartum management in women with VWD was of low quality. The ongoing high risk for PPH is evident, despite prophylactic treatment, as well as the need for higher quality evidence from larger prospective cohort studies to improve management strategies.

Acquired haemophilia A in a patient with breast cancer and lung carcinoma: a case report and literature review.

Acquired haemophilia A is a rare disorder caused by spontaneous formation of auto-antibodies (inhibitors) against coagulation factor VIII. This can lead tolife-threatening haemorrhages. Six to twenty-two percent of patients with acquired haemophilia have an underlying malignancy. We describe a 69-year-old woman with metastatic breast cancer and non-small cell lung carcinoma who presented at the emergency room with spontaneous bruising, and who was using a vitamin K antagonist. She had a prolonged activated partial thromboplastin time (aPTT) due to a coagulation factor VIII deficiency caused by factor VIII antibodies. She was treated with prednisone and cyclophosphamide.

Diagnosing and treating antiphospholipid syndrome: a consensus paper.

INTRODUCTION: The antiphospholipid syndrome (APS) is defined by the occurrence of venous and/or arterial thrombosis and/or pregnancy-related morbidity, combined with the presence of antiphospholipid antibodies (aPL) and/or a lupus anticoagulant (LAC). Large, controlled, intervention trials in APS are limited. This paper aims to provide clinicians with an expert consensus on the management of APS. METHODS: Relevant papers were identified by literature search. Statements on diagnostics and treatment were extracted. During two consensus meetings, statements were discussed, followed by a Delphi procedure. Subsequently, a final paper was written. RESULTS: Diagnosis of APS includes the combination of thrombotic events and presence of aPL. Risk stratification on an individual base remains challenging. 'Triple positive' patients have highest risk of recurrent thrombosis. aPL titres > 99th percentile should be considered positive. No gold standard exists for aPL testing; guidance on assay characteristics as formulated by the International Society on Thrombosis and Haemostasis should be followed. Treatment with vitamin K-antagonists (VKA) with INR 2.0-3.0 is first-line treatment for a first or recurrent APS-related venous thrombotic event. Patients with first arterial thrombosis should be treated with clopidogrel or VKA with target INR 2.0-3.0. Treatment with direct oral anticoagulants is not recommended. Patients with catastrophic APS, recurrent thrombotic events or recurrent pregnancy morbidity should be referred to an expert centre. CONCLUSION: This consensus paper fills the gap between evidence-based medicine and daily clinical practice for the care of APS patients.

Are serum autoantibodies associated with brain changes in systemic lupus erythematosus? MRI data from the Leiden NP-SLE cohort.

OBJECTIVE: The effect of serum autoantibodies on the brain of systemic lupus erythematosus (SLE) patients remains unclear. We investigated whether serum autoantibodies, individually and assessed in groups, are associated with specific brain-MRI abnormalities or whether these structural changes are associated with other SLE-related or traditional cardiovascular disease risk factors. METHODS: All patients underwent brain 3Tesla-MRI. White matter hyperintensities (WMHs), ischemic lesions, inflammatory-like lesions and cerebral atrophy were scored. Serum autoantibodies analyzed included lupus anticoagulant (LAC), anticardiolipine (aCL) IgG and IgM (first 3 also grouped into antiphospholipid autoantibodies (aPL)), anti-dsDNA, anti-SSA, anti-SSB, anti-RNP, and anti-Sm (the latter 5 grouped into SLE-related autoantibodies). Associations were assessed using logistic regression analysis adjusted for potential confounders. Furthermore, a sensitivity analysis including anti-Beta2 glycoprotein-1 antibodies (anti-ß2GP1) in the aPL group was performed and the potential modification role of the neuropsychiatric clinical status in the model was assessed. RESULTS: 325 patients (mean age 42 years (SD 14), 89% female) were included. The following MRI-brain abnormalities were found: WMHs (71%), lacunar infarcts (21%), gliosis (11%), micro-hemorrhages (5%), large hemorrhages (2%), inflammatory-like lesions (6%) and atrophy (14%). No associations were found between individual or total SLE-related autoantibodies and inflammatory-like lesions. A higher number of positive aPL was associated with lacunar infarcts (OR 1.37 (95%CI 1.02-1.99) and gliosis (OR 2.15 (1.37-3.37)). LAC was associated with lacunar infarcts in white matter (OR 3.38 (1.32-8.68)) and atrophy (OR 2.49 (1.01-6.15)), and aCL IgG with gliosis (OR 2.71 (1.05-7.02)). Among other variables, SLE patients with hypertension presented a higher chance for WMHs (OR 5.61 (2.52-12.48)) and lacunar infarcts in WM (OR 2.52 (1.10-5.74)) and basal ganglia (OR 8.34 (2.19-31.70)), while cumulative SLE-damage was correlated with lacunar infarcts in WM (OR 1.43 (1.07-1.90)), basal ganglia (OR 1.72 (1.18-2.51)) and cerebellum (OR 1.79 (1.33-2.41)). These associations were confirmed in the sensitivity analysis. CONCLUSIONS: Brain abnormalities in SLE represent different underlying pathogenic mechanisms. aPL are associated with ischemic brain changes in SLE, while the presence of SLE-related serum autoantibodies is not related to inflammatory-like lesions. Hypertension and cumulative SLE-damage associate with ischemic MRI-brain changes in SLE, suggesting the importance of accelerated atherosclerosis in this process.

Clinically relevant differences between assays for von Willebrand factor activity.

Essentials It is unclear whether there are differences between von Willebrand factor (VWF) activity assays. We compared the four most used VWF activity assays in 661 von Willebrand disease (VWD) patients. All assays correlated excellently, but a discrepant classification was seen in 20% of patients. Differences between VWF activity assays have a large impact on the classification of VWD. SUMMARY: Background Measuring the ability of von Willebrand factor (VWF) to bind to platelets is crucial for the diagnosis and classification of von Willebrand disease (VWD). Several assays that measure this VWF activity using different principles are available, but the clinical relevance of different assay principles is unclear. Objective To compare the four most widely used VWF activity assays in a large VWD patient population. Methods We measured VWF:RCo (ristocetin to activate VWF + whole platelets), VWF:GPIbR (ristocetin + platelet glycoprotein Ib receptor [GPIb] fragments), VWF:GPIbM (gain-of-function GPIb fragments that bind VWF spontaneously without ristocetin) and VWF:Ab (monoclonal antibody directed against the GPIb binding epitope of VWF to mimic platelets) in 661 VWD patients from the nationwide 'Willebrand in the Netherlands' (WiN) Study. Results All assays correlated excellently (Pearson r > 0.9), but discrepant results led to a different classification for up to one-fifth of VWD patients. VWF:RCo was not sensitive enough to classify 18% of patients and misclassified half of genotypic 2B VWD patients, especially those with p.Arg1306Trp. VWF:GPIbR was more sensitive, accurately classified the vast majority of patients, and was unaffected by the p.Asp1472His variant that causes artificially low VWF:RCo. VWF:GPIbM was the most precise assay but misclassified over a quarter of genotypic 2A, 2B and 3 patients. VWF:Ab, often not considered an actual VWF activity assay, performed at least equally to the other assays with regard to accurate VWD classification. Conclusion Although the different VWF activity assays are often considered similar, differences between assays have a large impact on the classification of VWD.

Correction of a dominant-negative von Willebrand factor multimerization defect by small interfering RNA-mediated allele-specific inhibition of mutant von Willebrand factor.

Essentials Substitution therapy for von Willebrand (VW) disease leaves mutant VW factor (VWF) unhindered. Presence of mutant VWF may negatively affect phenotypes despite treatment. Inhibition of VWF by allele-specific siRNAs targeting single-nucleotide polymorphisms is effective. Allele-specific inhibition of VWF p.Cys2773Ser improves multimerization. SUMMARY: Background Treatment of the bleeding disorder von Willebrand disease (VWD) focuses on increasing von Willebrand factor (VWF) levels by administration of desmopressin or VWF-containing concentrates. Both therapies leave the production of mutant VWF unhindered, which may have additional consequences, such as thrombocytopenia in patients with VWD type 2B, competition between mutant and normal VWF for platelet receptors, and the potential development of intestinal angiodysplasia. Most cases of VWD are caused by dominant-negative mutations in VWF, and we hypothesize that diminishing expression of mutant VWF positively affects VWD phenotypes. Objectives To investigate allele-specific inhibition of VWF by applying small interfering RNAs (siRNAs) targeting common single-nucleotide polymorphisms (SNPs) in VWF. This approach allows allele-specific knockdown irrespective of the mutations causing VWD. Methods Four SNPs with a high predicted heterozygosity within VWF were selected, and siRNAs were designed against both alleles of the four SNPs. siRNA efficiency, allele specificity and siRNA-mediated phenotypic improvements were determined in VWF-expressing HEK293 cells. Results Twelve siRNAs were able to efficiently inhibit single VWF alleles in HEK293 cells that stably produce VWF. Transient cotransfections of these siRNAs with two VWF alleles resulted in a clear preference for the targeted allele over the untargeted allele for 11 siRNAs. We also demonstrated siRNA-mediated phenotypic improvement of the VWF multimerization pattern of the VWD type 2A mutation VWF p.Cys2773Ser. Conclusions Allele-specific siRNAs are able to distinguish VWF alleles on the basis of one nucleotide variation, and are able to improve a severe multimerization defect caused by VWF p.Cys2773Ser. This holds promise for the therapeutic application of allele-specific siRNAs in dominant-negative VWD.

Analysis of current perioperative management with Haemate® P/Humate P® in von Willebrand disease: Identifying the need for personalized treatment.

INTRODUCTION: Patients with Von Willebrand disease (VWD) are regularly treated with VWF-containing concentrates in case of acute bleeding, trauma and dental or surgical procedures. AIM: In this multicentre retrospective study, current perioperative management with a von Willebrand factor (VWF)/Factor VIII (FVIII) concentrate (Haemate® P) in patients with VWD was evaluated. PATIENTS/METHODS: Patients with VWD undergoing minor or major surgery between 2000 and 2015, requiring treatment with a VWF/FVIII concentrate (Haemate® P), were included. Achieved VWF activity (VWF:Act) and FVIII during FVIII-based treatment regimens were compared to predefined target levels in national guidelines. RESULTS: In total, 103 patients with VWD (148 surgeries) were included: 54 type 1 (73 surgeries), 43 type 2 (67 surgeries) and 6 type 3 (8 surgeries). Overall, treatment resulted in high VWF:Act and FVIII levels, defined as ≥0.20 IU/mL above predefined levels. In patients with type 1 VWD, respectively, 65% and 91% of trough VWF:Act and FVIII levels were higher than target levels. In patients with type 2 and type 3 VWD, respectively, 53% and 57% of trough VWF:Act and 72% and 73% of trough FVIII levels were higher than target level. Furthermore, FVIII accumulation over time was observed, while VWF:Act showed a declining trend, leading to significantly higher levels of FVIII than VWF:Act. CONCLUSION: High VWF:Act and accumulation of FVIII were observed after perioperative FVIII-based replacement therapy in patients with VWD, both underlining the necessity of personalization of dosing regimens to optimize perioperative treatment.

Health-related quality of life, developmental milestones, and self-esteem in young adults with bleeding disorders.

BACKGROUND: The treatment of bleeding disorders improved in the last decades. However, the effect of growing up with bleeding disorders on developmental, emotional, and social aspects is understudied. Therefore, this study assesses HRQOL, developmental milestones, and self-esteem in Dutch young adults (YA) with bleeding disorders compared to peers. METHODS: Ninety-five YA (18-30 years) with bleeding disorders (78 men; mean 24.7 years, SD 3.5) and 17 women (mean 25.1 years, SD 3.8) participated and completed the Pediatric Quality of Life Inventory Young Adult version, the Course of Life Questionnaire, and the Rosenberg Self-Esteem Scale. Differences between patients with bleeding disorders and their peers, and between hemophilia severity groups, were tested using Mann-Whitney U tests. RESULTS: YA men with bleeding disorders report a slightly lower HRQOL on the total scale, physical functioning, and school/work functioning in comparison to healthy peers (small effect sizes). YA men with severe hemophilia report more problems on the physical functioning scale than non-severe hemophilia. YA men with bleeding disorders achieved more psychosexual developmental milestones than peers, but show a delay in 'paid jobs, during middle and/or high school.' A somewhat lower self-esteem was found in YA men with bleeding disorders in comparison to peers (small effect size). For YA women with bleeding disorders, no differences were found on any of the outcomes in comparison to peers. CONCLUSION: This study demonstrates some impairments in HRQOL and self-esteem in YA men with bleeding disorders. By monitoring HRQOL, problems can be identified early, especially with regard to their physical and professional/school functioning.

Comparison of haemostatic function of PAS-C-platelets vs. plasma-platelets in reconstituted whole blood using impedance aggregometry and thromboelastography.

BACKGROUND AND OBJECTIVES: There are concerns about the haemostatic function of platelets stored in platelet additive solution (PAS). Aim of this study was to compare the haemostatic function of PAS-C-platelets to plasma-platelets in reconstituted whole blood. MATERIALS AND METHODS: In our experiment, whole blood was reconstituted with red blood cells, solvent-detergent (SD) plasma and either PAS-C-platelets or plasma-platelets (n = 7) in a physiological ratio. On storage days 2, 5, 8 and 13, the agonist-induced aggregation (multiple electrode aggregometry), clot formation (thromboelastography) and agonist-induced CD62P responsiveness (flow cytometry) were measured. RESULTS: Samples with PAS-C-platelets showed significantly lower aggregation than plasma-platelets when induced with adenosine diphosphate, -6 U (95% confidence interval: -8; -4) or thrombin receptor-activating protein, -15 U (-19; -10). Also when activated with collagen and ristocetin, the PAS-C-platelets showed less aggregation, although not statistically significant. All samples with PAS-C-platelets showed significantly lower agonist-induced CD62P responsiveness than samples with plasma-platelets. However, there was no difference regarding all TEG parameters. CONCLUSION: Our findings demonstrate that the function - aggregation and CD62P responsiveness - of PAS-C-platelets in reconstituted whole blood is inferior to that of plasma-platelets, which may have implications in the setting of massive transfusions.

Plasma levels of plasminogen activator inhibitor-1 and bleeding phenotype in patients with von Willebrand disease.

INTRODUCTION: von Willebrand disease (VWD) is the most common inherited bleeding disorder. In VWD patients, large variations in bleeding tendency are observed, which cannot be completely explained by the variation in von Willebrand factor levels or activities. Thus, there must be additional factors, for instance, changes in fibrinolysis that have an effect on the variation in bleeding tendency in VWD patients. AIM: To investigate whether plasminogen activator inhibitor-1 (PAI-1) level influences the variation in bleeding tendency in VWD patients. METHODS: PAI-1 antigen levels were measured in the plasma of 633 patients with moderate or severe VWD who participated in the 'Willebrand in the Netherlands' (WiN) study, a nationwide multicentre cross-sectional study. Bleeding severity was assessed using the Tosetto bleeding score. RESULTS: PAI-1 levels increased with age (Spearman's rho: 0.225, P < 0.001) and were higher in men (23 [IQR 12-60] vs. 20 [IQR 10-44] ng mL-1 in women, P = 0.039), whereas the bleeding score was higher in women (11 [IQR 7-17] vs. 9 [IQR 5-14] ng mL-1 in men, P = 0.002). After adjustment for age and sex by stratification, PAI-1 level and bleeding score were negatively correlated (Spearman's rho: -0.170, P = 0.017) in the group of 196 young (age ≤ 45 year) female VWD patients, accounting for 31% of our study population. CONCLUSION: In young female VWD patients, we observed that low PAI-1 levels were associated with a higher bleeding score, which may partly explain the observed variability in bleeding phenotype in VWD patients.

State of the art: von Willebrand disease.

The State of the Art in von Willebrand disease (VWD) has been impacted not only by discoveries in the field of haemostasis, but also by changes in practice in other fields. The development of bleeding assessment tools has led to the clarification of bleeding symptoms and phenotype in VWD. New discoveries in the biology and genetics of von Willebrand factor (VWF) are challenging our existing diagnostics and classification(s). An improved understanding of reproductive physiology and the pathology of VWD along with changing obstetric, gynaecologic and haemostatic therapies necessitate an evolving response to the care of women with VWD. The survival of patients with autoimmune disease, malignancies and congenital heart disease along with increasing use of circulatory support devices and extracorporeal membrane oxygenation is increasing the prevalence of acquired von Willebrand syndrome. In each of these challenges, there are opportunities to improve the care of our patients with VWD.

Joint surgery in von Willebrand disease: a multicentre cross-sectional study.

BACKGROUND: Joint bleeds are reported by 23% of von Willebrand disease (VWD) patients and associated with orthopaedic surgery. Limited data are available on joint surgery in VWD. AIM: To assess the prevalence, indications, management and complications of joint surgery in VWD patients. METHODS: 804 VWD patients with historically lowest von Willebrand factor (VWF) activity ≤30 U dL-1 completed a questionnaire on joint bleeds, joint damage and orthopaedic surgery. We retrieved additional medical file data of patients who underwent surgery on large joints (shoulder, elbow, hip, knee or ankle). RESULTS: 116 out of 804 patients (14%) reported large joint surgery. Compared to VWD patients without previous orthopaedic surgery, these 116 patients reported more frequently a history of joint bleeds and joint damage (41% vs. 20%, P < 0.001 and 61% vs. 20%, P < 0.001). Medical file data on 126 large joint surgeries in 79 VWD patients revealed that this surgery was associated with joint damage due to prior joint bleeds in 24% of the procedures. Preoperative clotting factor correction (CFC) to prevent bleeding was administered in most cases (81%). Documentation on postoperative bleeding was found in 23 surgeries (18%). CONCLUSIONS: Large joint surgery is reported by 14% of VWD patients, related to joint bleeds in 24% and seems associated with bleeding complications frequently despite perioperative CFC.

Developments in the diagnostic procedures for von Willebrand disease.

Von Willebrand disease (VWD) is the most common inherited bleeding disorder but its diagnosis can be challenging due to the heterogeneity of the disease. VWD is mainly associated with mild mucocutaneous bleeding, although there are more severe phenotypes with bleeding from the gastrointestinal tract or even the joints. Also, surgical interventions and trauma may lead to critical bleeding events. These bleeding episodes are all related to quantitative or qualitative defects of von Willebrand factor (VWF), a multimeric glycoprotein produced by endothelial cells and megakaryocytes, which mediates platelet adhesion and aggregation and binds factor VIII (FVIII) in the circulation. This review describes the diagnostic procedures required for correct diagnosis. Accurate diagnosis and classification is required for proper treatment and counseling. Assessment of bleeding starts with the medical history. After a positive bleeding or family history, subsequent laboratory investigations will start with a panel of standard screening tests for hemostatic defects. Patients suspected of having VWD will be tested for plasma VWF antigen levels, the ability of VWF to bind platelets and FVIII activity. When VWD is confirmed, a set of subtyping tests can classify the patients as VWD types 1, 2 (A, B, M or N) or 3. The performance of some additional assays and analyses, such as VWF propeptide measurement or genetic analysis, may help in identifying the pathological mechanism behind certain defects or can guide in the choice of treatment.

Angiogenic characteristics of blood outgrowth endothelial cells from patients with von Willebrand disease.

BACKGROUND: Endothelial von Willebrand factor (VWF) inhibits angiogenesis. Accordingly, blood outgrowth endothelial cells (BOECs) isolated from von Willebrand disease (VWD) patients showed enhanced in vitro angiogenesis when compared with healthy control BOECs. Characterization of the angiogenic response of VWD BOECs is limited and differences between the different types of VWD have not been investigated in detail. OBJECTIVES: The aim of this study was to further explore the potential pathogenic effect of VWF mutations on angiogenesis. METHODS: BOECs were isolated from four healthy individuals, 10 patients with VWD and one heterozygous carrier of a type 2N mutation. Cell migration and tube formation were measured. RESULTS: Migration velocity and total tube formation were similar between VWD patients and controls in general. BOECs from the type 3 VWD patient and one type 2B patient showed increased migratory velocity and tube formation compared with BOECs from other patients and healthy controls. Directional migration was impaired in eight out of 10 VWD BOECs and the ability to form tubes was limited to early passage numbers, but not for BOECs from healthy controls. CONCLUSION: BOECs can be a useful tool for ex vivo assessment of endothelial cell function in patients with different types of VWD, but possible limitations, such as early loss of angiogenic capacity, should be recognized. BOECs from most VWD patients consistently showed impairment in the directionality of migration. This is the first report on angiogenic properties of a type 3 VWD BOEC, which showed increased in vitro angiogenesis.

CLEC4M and STXBP5 gene variations contribute to von Willebrand factor level variation in von Willebrand disease.

BACKGROUND: von Willebrand factor (VWF) levels in healthy individuals are influenced by variations in genetic loci other than the VWF gene, whose contribution to VWF levels in patients with von Willebrand disease (VWD) is largely unknown. OBJECTIVES: To investigate the association between single-nucleotide polymorphisms (SNPs), VWF levels, and bleeding phenotype. PATIENTS/METHODS: In 364 type 1 VWD and 240 type 2 VWD patients from the nationwide cross-sectional 'Willebrand in The Netherlands' (WiN) study, we studied the association between eight SNPs in STXBP5, SCARA5, ABO, VWF, STAB2, STX2, TC2N, and CLEC4M, and VWF antigen (VWF:Ag), VWF activity (VWF:Act), and bleeding phenotype as assessed with the Tosetto bleeding score. RESULTS: In type 1 patients, STXBP5 was associated with a lower VWF:Ag level (adjusted difference of -3.0 IU dL(-1) per allele; 95% confidence interval [CI] -6.0 to 0.1) and CLEC4M with both a lower VWF:Ag level (-4.3 IU dL(-1) per allele; 95% CI -7.9 to -0.6) and lower VWF:Act (-5.7 IU dL(-1) per allele; 95% CI -10.9 to -0.5). In type 2 patients, none of the SNPs was associated with VWF levels. None of the genetic variants was associated with bleeding score. CONCLUSIONS: Genetic variations in STXBP5 and CLEC4M are associated with VWF level variation in type 1 VWD, but not in type 2 VWD. This study increases our understanding of the pathophysiology of VWD, and provides a further indication of the involvement of STXBP5 and CLEC4M in determining VWF levels in VWD.